N2 -naphthalenesulfonyl-L-argininamides, and pharmaceutical salts, compositions and methods

ABSTRACT

N 2  -naphthalenesulfonyl-L-argininamides and the pharmaceutically acceptable salts thereof have been found to be effective as pharmaceutical agents for the inhibition and suppression of thrombosis.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation in part of applications Ser. No. 671,436 March 29, 1976, and Ser. No. 671,568 filed Mar. 29, 1976, now Pat. No. 4,049,645, issued Sept. 20, 1977, which in turn are divisionals of application Ser. No. 622,390 filed Oct. 14, 1975, now abandoned.

BACKGROUND OF THE INVENTION

1. Field of the Invention

This invention relates to the discovery of certain new and useful N² -naphthalenesulfonyl-L-argininamides and the pharmaceutically acceptable salts thereof, which are of especial value in view of their outstanding antithrombotic properties and low toxicities.

2. Description of the Prior Art

In the past, there have been many attempts to obtain new and imporved agents for the treatment of thrombosis. The N² -(p-tolylsulfonyl)-L-arginine esters have been found to be one type of agent which can be used and these have been found to be effective in dissolving blood clots. (U.S. Pat. No. 3,622,615, issued Nov. 23, 1971)

One family of compounds which have been found to be particularly useful as highly specific inhibitors of thrombin for the control of thrombosis is the N² -dansyl-L-arginine ester or amide. (Our pending U.S. application Ser. No. 496,939, filed Aug. 13, 1974 now U.S. Pat. No. 2,978,045)

However, there is a continuing need for a highly specific inhibitor on thrombin for the control of thrombosis, which exhibits lower toxicity.

SUMMARY OF THE INVENTION

It has now been discovered that N² -naphthalenesulfonyl-L-argininamides exhibit antithrombotic activity and even lower toxicity levels at the same relative potencies, as compared with the N² -dansyl-L-arginine ester or amide.

The compounds of this invention can be represented by the formula (I): ##STR1## wherein R₁ is selected from the group consisting of naphthyl, 5,6,7,8-tetrahydronaphthyl and naphythyl substituted with at least one substituent selected from the group consisting of halo, hydroxy, nitro, cyano, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxy and C₂ -C₂₀ dialkylamino; R₂ is selected from the group consisting of hydrogen, C₁ -C₁₀ alkyl, C₇ -C₁₂ aralkyl and C₆ -C₁₀ aryl; Z is oxy, thio or sulfinyl; an n is an integer of 0 or 1. Also encompassed within this invention are pharmaceutically acceptable salts thereof.

This invention also relates to a method for inhibiting activity and suppressing activation of thrombin in vivo, which comprises introducing into a living body a pharmaceutically effective amount of an N² -naphthalenesulfonyl-L-argininamide or the pharmaceutically acceptable salts thereof.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

This invention relates to a group of N² -naphthalenesulfonyl-L-argininamides of the formula (I): ##STR2## wherein R₁ is naphthyl, such as 1-naphthyl and 2-naphthyl, 5,6,7,8-tetrahydronaphthyl, such as 5,6,7,8-tetrahydro-1-naphthyl and 5,6,7,8-tetrahydro-2-naphthyl, or naphthyl substituted with at least one substituent selected from the group consisting of halo, such as fluoro, chloro, bromo and iodo, hydroxy, nitro, cyano, alkyl of 1-10 (preferably 1-5) carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl or the like, alkoxy of 1-10 (preferably 1-5) carbon atoms, such as methoxy, ethoxy, propoxy, isopropoxy, butoxy, sec-butoxy, tert-butoxy, pentyloxy or the like, and dialkylamino of 2-20 (preferably 2-10) carbon atoms, such as dimethylamino, diethylamino, N-methyl-N-ethylamino or the like; R₂ is hydrogen, alkyl of 1-10 (preferably 1-6) carbon atoms, such as methyl, ethyl, propyl, butyl, tert-butyl, hexyl, octyl, decyl or the like, C₆ -C₁₀ aryl such as phenyl or naphthyl, and preferably phenyl, or aralkyl of 7-12 (preferably 7-10) carbon atoms, such as benzyl, phenethyl or the like; Z is oxy (--0--), thio (--S--) or sulfinyl (--SO--); and n is an integer of 0 or 1.

Suitable illustrations of R₁ in the above formula (I) are 1-naphthyl, 2-naphthyl, 5,6,7,8-tetrahydro-1-naphthyl, 5,6,7,8-tetrahydro-2-naphthyl, 6-methyl-2-naphthyl, 6-methyl-1-naphthyl, 7-methyl-1-naphthyl, 7-methyl-2-naphthyl, 6-ethyl-2-naphthyl, 6,7-dimethyl-1-naphthyl, 6,7-dimethyl-2-naphthyl, 6-isopropyl-2-naphthyl, 5-chloro-1-naphthyl, 6-chloro-2-naphthyl, 6-bromo-1-naphthyl, 5-hydroxy-1-naphthyl, 7-hydroxy-2-naphthyl, 5-dimethylamino-1-naphthyl, 5-dimethylamino-2-naphthyl, 5-diethylamino-1-naphthyl, 6-dimethylamino-1-naphthyl, 6-dimethylamino-2-naphthyl, 5-methoxy-1-naphthyl, 6-methoxy-2-naphthyl, 7-methoxy-2-naphthyl, 4,6-dimethoxy-2-naphthyl, 6,7-dimethoxy-2-naphthyl and 6,7-diethoxy-2-naphthyl.

Suitable R₂ groups in the above formula (I) are hydrogen, ethyl and tert-butyl.

Suitable ##STR3## groups are 3-carboxy-4-morpholino, 3-carboxy-4-thiomorpholino, 1-oxo-3-carboxy-4-thiomorpholino and 4-carboxy-3-thiazolidinyl.

Illustrative of suitable N² -naphthalenesulfonyl-L-argininamides of sufficient activity are the following:

4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]-3-carboxythiomorpholine 1-oxide

4-[N² -(6,7-dimethoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

3-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiazolidine-4-carboxylic acid

Of the compounds of this invention, it will be understood that the following compounds are most preferred due to their high level of antithrombotic activity and low level of toxicity.

4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

4-[N² -(6,7-dimethoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

The pharmaceutically acceptable salts of the above compounds are of course also included within the scope of this invention.

The above compounds are intended only to illustrate the variety of structures which can be used in the process of this invention, and the above listing is not to be construed as limiting the scope of the invention.

For the preparation of the compounds of this invention, various methods can be employed depending upon the particular starting materials and/or intermediates involved.

Successful preparation of these compounds is possible by way of several synthetic routes which are outlined below.

(a) Condensation of an L-argininamide with a naphthalenesulfonyl halide

This process may be illustrated as follows: ##STR4## In the above formulas, R₁, R₂, Z and n are as defined herein above, and X is halogen.

The N² -naphthalenesulfonyl-L-argininamide (I) is prepared by the condensation of an L-argininamide (II) with a substantially equimolar amount of a naphthalenesulfonyl halide (III), preferably a chloride.

The condensation reaction is generally effected in a suitable reaction-inert solvent in the presence of an excess of a base, such as an organic base (triethylamine, pyridine) or a solution of an inorganic base (sodium hydroxide, potassium carbonate), at a temperature of 0° C to the boiling temperature of the solvent for a period of 10 minutes to 15 hours.

The preferred solvents for the condensation include benzene-diethyl ether, diethyl ether-water and dioxane water.

After the reaction is complete, the formed salt is extracted with water, and the solvent is removed by such standard means as evaporation under reduced pressure to give the N² -naphthalenesulfonyl-L-argininamide (I), which can be purified by trituration or recrystallization from a suitable solvent, such as diethyl ethertetrahydrofuran, diethyl ether-methanol and water-methanol, or may be chromatographed on silica gel.

The L-argininamides (II) starting materials required for the condensation reaction can be prepared by protecting the guanidino and α-amino groups of the arginine via nitration, acetylation, formylation, phthaloylation, trifluoroacetylation, p-methoxybenzyloxycarbonylation, benzoylation, benzyloxycarbonylation, tert-butoxycarbonylation or tritylation and then condensing the formed N^(G) -substituted-N² -substituted-L-arginine with a corresponding secondary amine by such a conventional process as the acid chloride method, azide method, mixed anhydride method, activated ester method or carbodiimide method, and thereafter selectively removing the protective group.

(b) Removal of the N^(G) -substituent from an N^(G) -substituted-N² -naphthalenesulfonyl-L-argininamide

This process may be illustrated as follows: ##STR5##

In the above formulas, R₁, R₂, Z, X and n are as defined herein above; Y" is a protective group for the amino group, such as benzyloxycarbonyl or tertbutoxycarbonyl; and Y and Y' are hydrogen and protective groups for the guanidino group, such as nitro, tosyl, trityl, oxycarbonyl or the like.

At least one of Y and Y' is a protective group for the guanidino group.

The N² -naphthalenesulfonyl-L-argininamide (I) is prepared by removing the N^(G) -substituent from an N^(G) -substituted-N² -naphthalenesulfonyl-L-argininamide (VIII) by means of acidolysis or hydrogenolysis.

The acidolysis is generally effected by contacting the N^(G) -substituted-N² -naphthalenesulfonyl-L-argininamide (VIII) and an excess of an acid such as hydrogen fluoride, hydrogen chloride, hydrogen bromide or trifluoroacetic acid, without a solvent or in a solvent, such as an ether (tetrahydrofuran, dioxane), an alcohol (methanol, ethanol) or acetic acid at a temperature of -10° C to 100° C, and preferably at room temperature for a period of 30 minutes to 24 hours.

The products are isolated by evaporation of the solvent and the excess acid, or by trituration with a suitable solvent followed by filtration and drying.

Because of the use of the excess acid, the products are generally the acid addition salts of the N² -naphthalenesulfonyl-L-argininamides (I), which can be easily converted to a free amide by neutralization.

The removal of the nitro group and the oxycarbonyl group, e.g., benzyloxycarbonyl, p-nitrobenzyloxycarbonyl, is readily accomplished by the hydrogenolysis.

The hydrogenolysis is effected in a reaction-inert solvent, e.g., methanol, ethanol, tetrahydrofuran or dioxane, in the presence of a hydrogen-activating catalyst, e.g., Raney nickel, palladium, or platinum, in a hydrogen atmosphere at a temperature of 0° C to the boiling temperature of the solvent for a period of 2 hours to 120 hours.

The hydrogen pressure is not critical, and atmospheric pressure is sufficient.

The N² -naphthalenesulfonyl-L-argininamides (I) are isolated by filtration of the catalyst followed by evaporation of the solvent.

The N² -naphthalenesulfonyl-L-argininamides can be purified in the same manner as described above.

The N^(G) -substituted-N² -naphthalenesulfonyl-L-argininamides (VIII) starting materials can be prepared by condensing an N^(G) -substituted-N² -substituted L-arginine (IV) (generally the N² -substituent is a protective group for the amino group, such as benzyloxycarbonyl, tert-butoxycarbonyl, or the like) and a corresponding secondary amine (V) via the azide method, mixed anhydride method, activated ester method, carbodiimido method or the like, selectively removing only the N² -substituent of an N^(G) -substituted-N² -substituted-L-arginamide (VI) by means of catalytic hydrogenolysis or acidolysis, and then condensing the thus obtained N^(G) -substituted-L-argininamide (VII) with a naphthalenesulfonyl halide (III), preferably a chloride in the presence of a base in a solvent. These reaction conditions are as described above in the condensation of an L-argininamide with a naphthalenesulfonyl halide, and the removal of the N^(G) -substituent from an N^(G) -subsituted-N² -naphthalenesulfonyl-L-argininamide.

(c) Condensation of an N² -naphthalenesulfonyl-L-arginyl halide with an amine

This process may be illustrated as follows: ##STR6##

In the above formulas, R₁, R₂, Z, X and n are as defined herein above.

The N² -naphthalenesulfonyl-L-argininamide (I) is prepared by the condensation of an N² -naphthalenesulfonyl-L-arginyl halide (IX), preferably a chloride with at least an equimolar amount of a secondary amine (V). The condensation reaction can be carried out without an added solvent. However, satisfactory results will be obtained with the use of a solvent such as basic solvents (dimethylformamide, dimethylacetamide, etc.) or halogenated solvents (chloroform, dichloromethane, etc.).

The amount of the solvent to be used is not critical and may vary from about 5 to 100 times the weight of the N² -naphthalenesulfonyl-L-arginyl halide (IX). Preferred condensation reaction temperatures are in the range of from -10° C to room temperature. The reaction time is not critical, but varies with the secondary amine (V) employed. In general, a period of from 5 minutes to 10 hours is operable.

The obtained N² -naphthalenesulfonyl-L-argininamide can be isolated and purified in the same manner as described above.

The N² -naphthalenesulfonyl-L-arginyl halide (IX) starting materials required for the condensation reaction can be prepared by reacting an N² -naphthalenesulfonyl-L-arginine with at least an equimolar amount of a halogenating agent such as thionyl chloride, phosphorous oxychloride, phosphorus trichloride, phosphorous pentachloride or phosphorus tribromide. The halogenation can be carried out with or without an added solvent. The preferred solvents are chlorinated hydrocarbons such as chloroform and dichloromethane, and ethers such as tetrahydrofuran and dioxane.

The amount of the solvent to be used is not critical and may vary from about 5 to 100 times the weight of the N² -naphthalenesulfonyl-L-arginine.

Preferred reaction temperatures are in the range of -10° C to room temperature. The reaction time is not critical, but varies with the halogenating agent and reaction temperature. In general, a period of 15 minutes to 5 hours is operable.

(d) Guanidylation of an N² -naphthalenesulfonyl-L-ornithinamide or an acid addition salt thereof

This process may be illustrated as follows: ##STR7##

In the above formulas, R₁, R₂, Z and n are as defined herein above.

The N² -naphthalenesulfonyl-L-argininamide (I) is prepared by guanidylating an N² -naphthalenesulfonyl-L-ornithinamide (X) with an ordinary guanidylating agent such as an O-alkylisourea, S-alkylisothiourea, 1-guanyl-3,5-dimethylpyrazole or carbodiimide. The preferred guanidylating agents are the O-alkylisourea and the S-alkylisothiourea.

The guanidylation of the N² -naphthalenesulfonyl-L-ornithinamide (X) with the O-alkylisourea or S-alkylisothiourea is generally effected in a solvent in the presence of a base at a temperature of from 0° C to the boiling temperature of the solvent for a period of from 30 minutes to 50 hours.

Examples of the preferred bases are triethylamine, pyridine, sodium hydroxide and sodium methoxide. The base is used in an amount of 0.01 to 0.1 equivalent to the N² -naphthalenesulfonyl-L-ornithinamide. Examples of the preferred solvents are water, water-ethanol and water-dioxane.

After the reaction is complete, the N² -naphthalenesulfonyl-L-argininamide (I) is isolated by evaporation of the solvent followed by removal of the excess base and the formed salt by a water wash.

It is well recognized in the art that an ester derivative of the N² -naphthalenesulfonyl-L-argininamide (I) wherein R₂ is alkyl, aralkyl or aryl, can be prepared from a carboxylic acid derivative of the N² -naphthalenesulfonyl-L-argininamide wherein R₂ is hydrogen, by the conventional esterification methods well known to those skilled in the art. It is also well recognized in the art that the carboxylic acid derivative can be prepared from the ester derivative by the conventional hydrolysis or acidolysis methods. The conditions under which esterification, hydrolysis or acidolysis would be carried out will be each apparent to those skilled in the art.

The N² -naphthalenesulfonyl-L-argininamide (I) of this invention forms acid addition salts with any of a variety of inorganic and organic acids. Some of the N² -naphthalenesulfonyl-L-argininamide containing a free carboxy group, wherein R₂ is hydrogen, forms salts with any of a variety of inorganic and organic bases.

The product of the reactions described above can be isolated in the free form or in the form of salts. In addition, the product can be obtained as pharmaceutically acceptable acid addition salts by reacting one of the free bases with an acid, such as hydrochloric, hydrobromic, hydroiodic, nitric, sulfuric, phosphoric, acetic, citric, maleic, succinic, lactic, tartaric, gluconic, benzoic, methanesulfonic, ethanesulfonic, benzenesulfonic, p-toluenesulfonic acid or the like. In a similar manner, the product can be obtained as pharmaceutically acceptable salts by reacting one of the free carboxylic acids with a base, such as sodium hydroxide, potassium hydroxide, ammonium hydroxide, triethylamine, procaine, dibenzylamine, 1-ephenamine, N,N'-dibenzylethylene-diamine, N-ethylpiperidine or the like.

Likewise, treatment of the salts with a base or acid results in a regeneration of the free amide.

As stated above, the N² -naphthalenesulfonyl-L-argininamides, and the salts thereof of this invention are characterized by highly specific inhibitory acitivty against thrombin as well as by their substantial lack of toxicity, and therefore these compounds are useful in the determination of thrombin in blood as diagnostic reagents, and/or for the medical control or prevention of thrombosis.

The antithrombotic activity of the N² -naphthalenesulfonyl-L-argininamide of this invention was compared with that of a known antithrombotic agent, N² -(p-tolylsulfonyl)-L-arginine methyl ester, by determining the fibrinogen coagulation time. The measurement of the fibrinogen coagulation time was conducted as follows:

An 0.8 ml aliquot of a fibrinogen solution, which had been prepared by dissolving 150 mg of bovine fibrinogen (Cohn fraction I) supplied by Armour Inc. in 40 ml of a borate saline buffer (pH 7.4), was mixed with 0.1 ml of a borate saline buffer, pH 7.4, (control) or a sample solution in the same buffer, and 0.1 ml of a thrombin solution (5 units/ml) supplied by Mochida Pharmaceutical Co., Ltd. was added to the solutions in an ice bath.

Immediately after mixing, the reaction mixture was transferred from the ice bath to a bath maintained at 25° C. Coagulation times were taken as the period between the time of transference to the 25° C bath and the time of the first appearance of fibrin theads. In the cases where no drug samples were added, the coagulation time was 50-55 seconds. The experimental results are summarized in Table 1. The term "concentration required to prolong the coagulation time by a factor of two" is the concentration of an active ingredient required to prolong the normal coagulation time 50-55 seconds to 100-110 seconds.

The concentration required to prolong the coagulation time by a factor of two for the known antithrombotic agent, N² -(p-tolylsulfonyl)-L-arginine methyl ester, was 1,100 μm. The inhibitors are shown in Table 1 by indicating R₁, R₂, Z and n in the formula (I) and the addition moiety. When a solution containing an N² -naphthalenesulfonyl-L-argininamide of this invention was administered intravenously into animal bodies, the high antithrombotic activity in the circulating blood was maintained for from 1 to 3 hours. The halflife for decay of the anti-thrombotic compounds of this invention in circulating blood was shown to be approximately 60 minutes; the physiological conditions of the host animals (rat, rabbit, dog and chimpanzee) were well maintained. The experimental decrease of fibrinogen in animals caused by infusion of thrombin was satisfactorily controlled by simultaneous infusion of the compounds of this invention.

The acute toxicity values (LD₅₀) determined by intraperitoneal administration of substances of formula (I) in mice (male, 20 g) range from about 1,000 to 10,000 milligrams per kilogram of body weight. A representative LD₅₀ value, for example, for 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid is more than 1,000 milligrams per kilogram. On the other hand, LD₅₀ values for N² -dansyl-N-butyl-L-argininamide and N² -dansyl-N-methyl-N-butyl-L-argininamide are 75 and 70 milligrams per kilogram, respectively.

The therapeutic agents of this invention may be administered alone or in combination with pharmaceutically acceptable carriers, the proportion of which is determined by the solubility and chemical nature of the compound, chosen route of administration and standard pharmaceutical practice. For example, the compounds may be injected parenterally, that is, intramuscularly, intravenously or subcutaneously. For parenteral administration, the compounds may be used in the form of sterile solutions containing other solutes, for example, sufficient saline or glucose to make the solution isotonic. The compounds may be administered orally in the form of tablets, capsules, or granules containing suitable excipients such as starch, lactose, white sugar and the like. The compounds may be administered sublingually in the form of troches or lozenges in which each active ingredient is mixed with sugar or corn syrups, flavoring agents and dyes, and then dehydrated sufficiently to make the mixture suitable for pressing into solid form. The compounds may be administered orally in the form of solutions which may contain coloring and flavoring agents. Physicians will determine the dosage of the present therapeutic agents which will be most suitable, and dosages vary with the mode of administration and the particular compound chosen. In addition, the dosage will vary with the particular patient under treatment. When the composition is administered orally, a larger quantity of the active agent will be required to produce the same effect as caused with a smaller quantity given parenterally. The therapeutic dosage is generally 10-50 mg/kg of active ingredient parenterally, 10-500 mg/kg orally per day. Having generally described the invention, a more complete understanding can be obtained by reference to certain specific examples, which are included for purposes of illustration only and are not intended to be limiting unless otherwise specified.

EXAMPLE 1

(a) n² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl chloride hydrochloride

A suspension of 2.5 g of N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginine in 10 ml of thionyl chloride containing a few drops of dimethyl formamide was stirred for 2 hours at room temperature.

Addition of cold dry diethyl ether resulted in a precipitate which was collected by filtration and washed several times with dry diethyl ether to give N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl chloride hydrochloride.

(B) Ethyl 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylate

To a stirred solution of 1.6 g of ethyl thiomorpholine-3-carboxylate and 3 ml of triethylamine in 40 ml of chloroform, which was cooled in an ice bath, there was carefully added N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl chloride hydrochloride obtained above. The reaction mixture was allowed to stand overnight at room temperature.

The reaction mixture was washed twice with 20 ml of saturated sodium chloride solution and evaporated to dryness. The residue was dissolved in water-ethanol and adjusted to pH 12 with 5N NaOH at a temperature below 0° C to precipitate a solid. This was collected by filtration to give 2.3 g of ethyl 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylate.

I.R. (KBr): 3,400 (broad), 1,735, 1,640, 1,160 cm⁻¹ Analysis -- Calcd. for C₂₄ H₃₃ O₆ N₅ S₂ (percent): C, 52.25; H, 6.03; N, 12.70 Found (percent): C, 52.36; H, 5.98; N, 12.51

(c) 4-[n² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

A solution of 2 g of ethyl 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylate in 15 ml of methanol and 4 ml of 2N sodium hydroxide solution was warmed to 50° C and held at that temperature for 24 hours. At the end of this period, the reaction mixture was concentrated and chromatographed on 200 ml of Daiaion® SK 102 ion exchange resin (200-300 mesh, H⁺ form, manufactured by Mitsubishi Chemical Industries Limited) packed in water, washed with ethanol-water (1:4) and eluted with ethanol-water-NH₄ OH (10:9:1). The main fraction was evaporated to dryness and washed with ether to give 1.3 g of 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid.

I.R. (KBr): 3,380 (broad), 1,620, 1,380, 1,155 cm⁻¹ Analysis -- Calcd. for C₂₂ H₂₉ O₆ N₅ S₂ (percent): C, 50.46; H, 5.58; N, 13.38 Found (percent): C, 50.61; H, 5.63; N, 13.40

EXAMPLE 2

(a) ethyl 4-[N^(G) -nitro-N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylate

To a well stirred solution of 4 g of ethyl 4-(N^(G) -nitro-L-arginyl)morpholine-3-carboxylate hydrochloride and 2 ml of triethylamine in 40 ml of chloroform, there was added in portions 1.5 g of 7-methoxy-2-naphthalenesulfonyl chloride.

The reaction mixture was stirred overnight at room temperature. At the end of this period, the chloroform solution was washed consecutively with 10% citric acid, saturated NaCl, saturated NaHCO₃ and saturated NaCl solutions. The chloroform solution was evaporated and the residue was chromatographed on silica gel packed in chloroform, and eluted from chloroform containing 3% ethanol. The main fraction was evaporated to dryness to give 1.6 g of ethyl 4-[N^(G) -nitro-N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylate

(B) 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]-morpholine-3-carboxylic acid

To a solution of 1.2 g of ethyl 4-[N^(G) -nitro-N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylate in 20 ml of tert-butanol and 4 ml of acetic acid, there was added 0.5 g of palladium black, and then the mixture was shaken in a hydrogen atmosphere for 48 hours at room temperature. The solution was filtered to remove the catalyst and evaporated to give an oily product. This was dissolved in 10 ml of methanol and 5 ml of 2N NaOH solution, warmed at 50° C for 24 hours and then evaporated to dryness. The residue was chromatographed on 300 ml of Diaion®SK102 ion exchange resin (100-200 mesh, H⁺ form, manufactured by Mitsubishi Chemical Industries Limited) packed in water, washed with 50% ethanol-water and eluted with ethanol-water-NH₄ OH (5:4:1). The main fraction eluted from the ethanol-water-NH₄ OH solution was evaporated to dryness to give a gummy material. This was recrystallized from ethanol-water to give 0.5 g of 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid in the form of a powder.

I.R. (KBr): 3,320 (broad), 1,620, 1,390, 1,155 cm⁻¹ Analysis-Calcd. for C₂₂ H₂₉ O₇ N₅ S₁ (percent): C, 52.06; H, 5.76; N, 13.80 Found (percent): C, 52.31, H, 5.81, N, 13.61

The following compounds are prepared in accordance with the procedure of the above examples.

4-[N² -(7-chloro-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

4-[N² -(7-hydroxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

4-[N² -(5-nitro-1-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

4-[N² -(5-cyano-1-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

4-[N² -(5-methoxy-1-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

Ethyl 4-[N² -(4,6-dimethoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylate

4-[N² -(5-ethoxy-1-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

4-[N² -(5-dimethylamino-1-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

3-[N² -(1-naphthalenesulfonyl)-L-arginyl]thiazolidine-4-carboxylic acid

4-[N² -(5,6,7,8-tetrahydro-2-naphthalenesulfonyl)-L-arginyl]-3-carboxythiomorpholine 1-oxide

4-[N² -(7-methyl-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

Ethyl 4-[N² -(7-methyl-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylate

4-[N² -(7-methyl-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid

4-[N² -(6,7-dimethyl-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid

Various other N² -naphthalenesulfonyl-L-argininamides or acid addition salts thereof were synthesized in accordance with the procedure of the above examples, and the test results are summarized in Table 1.

                                      TABLE 1                                      __________________________________________________________________________     Compound                                                                        ##STR8##                      Concentration required to prolong                                                       Prep- aration                                                                           Elementary analysis           Sam-                      Addi-                                                                              coagulation                                                                            process  Upper:Calculated                ple                       tion                                                                               time by a                                                                              (Ex.                                                                               m.p. Lower:Found                                                                             I.R. (KBr)             No.                                                                               R.sub.1    R.sub.2                                                                             Z    n moiety                                                                             factor of two                                                                          No.)                                                                               (° C)                                                                        C  H  N  (cm.sup.-1)            __________________________________________________________________________         ##STR9##  C.sub.2 H.sub.5                                                                     S    1 --          1   powder                                                                              52.25  52.36                                                                      6.03 5.98                                                                         12.70 12.51                                                                       3,400, 1,735,                                                                  1,640, 1,160           2                                                                                  ##STR10## H    S    1 --          1   "    50.46 50.61                                                                       5.58 5.63                                                                         13.38 13.40                                                                       3,380, 1,620,                                                                  1,380, 1,155           3                                                                                  ##STR11## H    O    1 --  2μM  2   "    52.06 52.31                                                                       5.76 5.81                                                                         13.80 13.61                                                                       3,320, 1,620,                                                                  1,390, 1,155           4                                                                                  ##STR12## H    SO   1 --          1   "    48.96 49.13                                                                       5.42 5.38                                                                         12.98 12.75                                                                       3,350, 1,620,                                                                  1,380, 1,150           5                                                                                  ##STR13## H    O    1 --  5μM  1   "    51.38 51.45                                                                       5.81 5.86                                                                         13.03 13.12                                                                       3,350, 1,630,                                                                  1,255, 1,150           6                                                                                  ##STR14## H    S    0 --          1   "    49.50 49.31                                                                       5.34 5.40                                                                         13.75 13.68                                                                       3,350, 3,200,                                                                  1,622                  __________________________________________________________________________

EXAMPLE 3

Tablets suitable for oral administration

Tablets containing the ingredients indicated below may be prepared by conventional techniques.

    ______________________________________                                                                Amount per tablet                                       Ingredient             (mg)                                                    ______________________________________                                         4-[N.sup.2 -(7-methoxy-2-naphthalenesulfonyl)-                                                        250                                                     L-arginyl]morpholine-3-carboxylic acid                                         Lactose                140                                                     Corn starch             35                                                     Talcum                  20                                                     Magnesium stearate      5                                                      Total                     450 mg                                               ______________________________________                                    

EXAMPLE 4

Capsules for oral administration

Capsules of the below were made up by thoroughly mixing together batches of the ingredients and filling hard gelatin capsules with the mixture.

    ______________________________________                                                                Amount per capsule                                      Ingredient             (mg)                                                    ______________________________________                                         4-[N.sup.2 -(7-methoxy-2-naphthalenesulfonyl)-                                                        250                                                     L-arginyl]morpholine-3-carboxylic acid                                         Lactose                250                                                     Total                     500 mg                                               ______________________________________                                    

EXAMPLE 5

Sterile solution for infusion

The following ingredients are dissolved in water for intravenous perfusion and the resulting solution is then sterilized.

    ______________________________________                                         Ingredients             Amount (g)                                             ______________________________________                                         4-[N.sup.2 -(7-methoxy-2-naphthalenesulfonyl)-                                                         25                                                     L-arginyl]morpholine-3-carboxylic acid                                         Buffer system           As desired                                             Glucose                 25                                                     Distilled water         500                                                    ______________________________________                                    

Having now fully described the invention, it will be apparent to one of ordinary skill in the art that many changes and modifications can be made thereto without departing from the spirit of the invention as set forth herein. 

What is claimed as new and intended to be covered by letters patent is:
 1. N² -naphthalenesulfonyl-L-argininamides having the formula ##STR15## and the pharmaceutically acceptable salts thereof, wherein R₁ is selected from the group consisting of naphthyl, 5,6,7,8-tetrahydronaphth-1-yl, 5,6,7,8-tetrahydronaphth-2-yl and naphthyl substituted with one or two substituents selected from the group consisting of halo, nitro, cyano, hydroxy, C₁ -C₁₀ alkyl, C₁ -C₁₀ alkoxy, and di(C₁ -C₁₀) alkylamino; R₂ is selected from the group consisting of hydrogen, C₁ -C₁₀ alkyl, phenyl, naphthyl, phenyl-C₁ -C₆ -alkyl, and naphthyl-C₁ -C₂ alkyl; Z is oxy, thio, or sulfinyl; and n is an integer of 0 or
 1. 2. The compound of claim 1, wherein R₁ is selected from the group consisting of 1-naphthyl, 2-naphthyl, 5,6,7,8-tetrahydro-1-naphthyl, 5,6,7,8-tetrahydro-2-naphthyl, 6-methyl-2-naphthyl, 6-methyl-1-naphthyl, 7-methyl-1-naphthyl, 7-methyl-2-naphthyl, 6-ethyl-2-naphthyl, 6,7-dimethyl-1-naphthyl, 6,7-dimethyl-2-naphthyl, 6-isopropyl-2-naphthyl, 5-chloro-1-naphthyl, 6-chloro-2-naphthyl, 6-bromo-1-naphthyl, 5-hydroxy-1-naphthyl, 7-hydroxy-2-naphthyl, 5-dimethylamino-1-naphthyl, 5-dimethylamino-2-naphthyl, 5-diethylamino-1-naphthyl, 6-dimethylamino-1-naphthyl and 6-dimethylamino-2-naphthyl, 5-methoxy-1-naphthyl, 6-methoxy-2-naphthyl, 7-methoxy-2-naphthyl, 4,6-dimethoxy-2-naphthyl, 6,7-dimethoxy-2-naphthyl and 6,7-diethoxy-2-naphthyl; and R₂ is hydrogen, ethyl or tert-butyl; Z is oxy, thio or sulfinyl; and n is an integer of 0 or
 1. 3. The compound of claim 2, wherein Z is thio and n is
 0. 4. The compound of claim 2, wherein Z is oxy, thio or sulfinyl, and n is
 1. 5. A compound of claim 4, which is 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]thiomorpholine-3-carboxylic acid.
 6. A compound of claim 4, which is 4-[N² -(6,7-dimethoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid.
 7. A compound of claim 4, which is 4-[N² -(7-methoxy-2-naphthalenesulfonyl)-L-arginyl]morpholine-3-carboxylic acid.
 8. A pharmaceutical composition for inhibiting activity and suppressing activation of thrombin in vivo, which comprises an amount effective to inhibit activity and suppress activation of thrombin in vivo, of a compound of claim 1 and a pharmaceutically acceptable carrier.
 9. A method of inhibiting activity and suppressing activation of thrombin in vivo, which comprises administering to a patient an amount effective to inhibit activity and suppress activation of thrombin in vivo, of the N² -naphthalenesulfonyl-L-argininamide of claim
 1. 